Human semen contains cationic amyloid fibrils, termed the "Semen Enhancer of Virus Infection" (SEVI), which strongly enhance HIV-1 infection and may play an important role in viral transmission. Our preliminary data show that amyloid-binding molecules bind to SEVI, and block semen-mediated enhancement of HIV-1 infection. This suggests that (i) SEVI is responsible for semen-mediated enhancement of HIV infection and (ii) SEVI represents a novel microbicide target. We therefore propose to explore a novel, innovative approach to HIV-1 microbicide development, using agents that selectively target SEVI. This high-risk/high-reward approach is fundamentally different from traditional microbicidal strategies that target the virus itself, and is expected to be highly complementary with direct antiviral approaches. Indeed, our long-term goal is to use SEVI-targeting agents in combination with traditional microbicides, to achieve optimal antiviral effectiveness. In the R21 phase, we will test whether novel amyloid-binding small molecules inhibit semen-mediated enhancement of HIV infection. The feasibility of this approach has been established using two amyloid-binding small molecules which contain "shielding" oligo-ethylene glycol (EG) moieties: BTA-EG4 and -EG6. These agents efficiently inhibit SEVI- and semen-mediated enhancement of HIV infection. In Aim 1, we will generate and test novel derivatives of these and other amyloid-binding molecules, including oligovalent molecules that are expected to possess increased SEVI binding affinity. We will then test their ability to inhibit SEVI- and semen- mediated enhancement of HIV infection using a panel of R5 virus strains (including different clades and transmitted strains). In Aim 2, we will examine the interaction between novel amyloid-binding small molecules and cells from the female reproductive tract. We will evaluate whether our compounds are toxic to human cervicovaginal epithelial cells (HCEC), and we will test whether they inhibit SEVI-enhanced binding of HIV-1 to HCEC and/or SEVI-enhanced trans-infection of PBMC by HCEC exposed to HIV-1 virions. The R33 phase will be undertaken only if well-defined milestones are achieved. In Aim 3, we will use structure-activity relationship (SAR) data to refine our chemical compositions. We will also test whether our lead molecules have efficacy in a cervical explant model for HIV-1 infection, and whether they have a synergistic or additive effect on the ability of other candidate microbicides to inhibit HIV-1 infection in the presence of semen. In the final Aim, we will assess the toxicity and inflammatory effects of the most promising candidate molecules, using beneficial Lactobacillus strains and cervical explants. The R33 phase will culminate with an evaluation of the safety and tolerability of the most promising compound in the rabbit vaginal irritation (RVI) model. The overall goal of these studies is to carefully determine whether small molecules that target SEVI have potential utility as a novel class of microbicides. PUBLIC HEALTH RELEVANCE: New approaches to prevent the transmission of human immunodeficiency virus type-1 (HIV-1) are urgently needed. This application seeks to develop a new class of microbicidal agents that are targeted not to the virus itself, but to a host protein found in semen that strongly enhances HIV-1 infection. This high risk, high reward approach is fundamentally different from traditional microbicidal strategies that target the virus, and is expected to be highly complementary with direct antiviral approaches.